Axon degeneration occurs frequently in many types of chronic neurodegenerative diseases and in injuries to axons caused by toxic, ischemic, or traumatic insults. It may lead to separation of the neuron from its targets, resulting in loss of neuronal function. One model of axon degeneration is the self-destructive process observed at the distal portion of a transected axon upon injury, termed Wallerian degeneration (WD) as first described by Waller (1850). In the process of WD, if a nerve fiber is cut or crushed, the part distal to the injury (i.e. the part of the axon separated from the neuron's cell nucleus) will degenerate. Because most neuronal proteins are synthesised in the soma and carried to the axon by specialised axonal transport systems, degeneration of the transected axons has long been thought to result from starvation of necessary proteins and other materials. However, the discovery of a spontaneously occurring mutant mouse strain, C57BL/Wlds, whose axons survived for as long as weeks after transection suggested that Wallerian degeneration involves an active and regulated auto-destruction program.
Indeed one of the most striking cellular responses during WD in the peripheral nervous system (PNS) is the proliferation and infiltration of macrophages (Bruck, 1997). Macrophages participate in a wide array of cellular responses during WD. Once activated, they release factors that are mitogenic for Schwann cells (Baichwal et al., 1988). The completion of WD relies on the phagocytic ability of macrophages to degrade myelin and axonal debris (Griffin et al., 1992). In addition, macrophages can degrade molecules inhibitory to axonal regeneration (Bedi et al., 1992) as well as release factors, such as interleukin-1 (IL-1), which can promote axonal growth via the induction of neurotrophic factors such as nerve growth factor (NGF) (Lindholm et al., 1987).
The precise mechanisms responsible for macrophage recruitment during WD are not completely understood. One group of factors that may play a role in macrophage recruitment and activation is the serum complement proteins. The importance of complement proteins immune-mediated peripheral nerve injury has been investigated previously.
Mead et al. (2002) showed that C6 deficient PVG/c rats, unable to form the membrane attack complex (MAC), exhibit neither demyelination nor axonal damage and significantly reduced clinical score in the antibody-mediated experimental autoimmune encephalomyelitis (EAE) model for multiple sclerosis when compared with matched C6 sufficient rats. However, levels of mononuclear cell infiltration were equivalent to those seen in C6 sufficient rats. Mead et al. (2002) concluded that demyelination and axonal damage occur in the presence of Ab and require activation of the entire complement cascade, including MAC deposition.
Jung et al. (1995) disclosed that treatment with recombinant human soluble complement receptor type 1 (sCR1) markedly suppressed clinical signs of myelin-induced experimental autoimmune neuritis (EAN) in Lewis rats (an animal model of the human Guillain-Barré syndrome). Extended demyelination and axonal degeneration were also prevented. These findings underscore the functional importance of complement during inflammatory demyelination in the peripheral nervous system.
Indeed, in EAN, complement depletion diminished myelin breakdown and macrophage recruitment in vivo (Feasby et al., 1987; Vriesendorp et al., 1995). Other groups have suggested that inhibition of the complement cascade reduces damage in neurodegenerative disease of the central nervous system (CNS) (e.g. Woodruff et al. 2006; Leinhase et al. 2006).
Daily et al. (1998) disclose a significant reduction in the recruitment of macrophages into distal degenerating nerve in complement-depleted animals. Complement depletion also decreased macrophage activation, as indicated by their failure to become large and multivacuolated and their reduced capacity to clear myelin. In the normal situation the myelin is cleared, the proximal part of the nerve forms sprouts which slowly grow along the path of the degenerated nerve. However, regeneration is slow (2-2.5 mm/day) and the environment of a degenerated nerve is full of factors which inhibit the growth of the axon and the necessary growth factors can be limiting or even absent. Myelin itself has been proposed to be a major inhibiting factor. Therefore rapid clearance of myelin is considered a conditio sine qua non for axonal regeneration. Thus the delayed clearance of myelin in complement-depleted animals is expected to result in impaired axonal regeneration. These findings indicate a role for serum complement in both the recruitment and activation of macrophages during peripheral nerve degeneration as well as an active role for macrophages in promoting axonal regeneration.
Indeed U.S. Pat. No. 6,267,955 discloses the methods wherein mononuclear phagocytes are administered at or near a site of injury or disease of the central or peripheral nervous system of a mammal in order to effect removal of the myelin debris that reportedly inhibits axonal regeneration, and for release of macrophage-derived cytokines that promote modulation of astrocytes and oligodendrocytes so as to support axonal regeneration.
Axonal degeneration is the main cause of disability both in hereditary and in acquired demyelinating neuropathies. While most current therapeutic research aims at restoring myelination, the present inventors focus on the consequence of demyelination: secondary axonal degeneration. As a model we have used acute demyelination and axonal degeneration after crush injury and subsequent regeneration of the nerve. It is an object of the present invention to provide for means and methods that promote and improve regeneration of nerves.